PI: Gartenberg, Marc R. PROJECT SUMMARY Cohesin organizes eukaryotic genomes into structures that segregate faithfully between dividing cells. The complex keeps newly replicated sister chromatids together and folds individual chromatids into compact chromosomal structures. Cohesin accomplishes these tasks by holding distant DNA sites in close proximity. Defects in cohesin or cohesin regulators have been linked to cancer and are the cause of developmental diseases known collectively as cohesinopathies. Thus, understanding how cohesin functions will reveal much about chromosome structure as it relates to human health. Cohesin complexes bind numerous chromosomal sites by encircling the DNA of each site in a topological embrace. Many cohesin binding sites lie in and around genes. When the genes are expressed, cohesin must move out of the way of advancing RNA polymerases. How this occurs is not clear. The central goal of this proposal is to understand how cohesin moves on chromosomal arms to adopt positions that achieve structural roles yet permit proper gene expression. The Gartenberg lab recently showed that cohesin translocates on DNA by sliding, and that the complex remains cohesive while in motion. Using yeast as a model system, this proposal aims to determine how the dynamic distribution of cohesin on chromosomal DNA is determined by binding, sliding and function of the complex. Aim 1 uses novel, molecular biology assays to define the dimensions of the DNA channel through cohesin, as well as determine whether the complex embraces one or two chromatids within the same channel. The experiments will determine how the dimensions of the complex limit which obstacles the complex can slide past. Aim 2 uses molecular biology assays to determine how transcription and other ATP-dependent processes regulate cohesin movement, and define the biological consequences when movement is blocked. The experiments will determine the molecular basis and physiological benefit of cohesin accumulating at specific cohesin enrichment sites. Aim 3 uses genome-scale strategies to distinguish between chromosomal sites where cohesin holds sister chromatids together and where it does not. The experiments will yield the first comprehensive maps of cohesin based on the functionality of the complex.